Chromosomal aneuploidy is associated with a large number of genetic disorders that could be prevented or prepared for by appropriate diagnosis, e.g. Verp et al, chapter 7, in Filkins and Russo, Eds., Human Prenatal Diagnosis (1990). Such disorders include Down's syndrome associated with chromosome 21 trisomy, Edward's syndrome associated with chromosome 18 trisomy, Plateau's syndrome associated with chromosome 13 trisomy, Turner's syndrome associated with an absence of an X chromosome (XO), Kleinfelter's syndrome associated with an extra X chromosome (XXY), XYY syndrome, triple X syndrome, and the like. Presently, of the 15,600 annual births in the United States with chromosomal abnormalities (and 643,000 worldwide), 53% have sex chromosome aneuploidies, 26% have autosomal trisomies, and the remaining have autosomal rearrangements. There is a clear need for accurate, inexpensive, and convenient aneuploidy diagnostics.
A variety of diagnostic approaches have been taken or have been proposed to detect the presence of aneuploidy, e.g. construction and microscopic examination of karyotypes, flow cytometric examination of chromosomes, assay for enzymatic markers, in situ hybridization, quantitative amplification of selected chromosome-specific markers, and the like, e.g. Khan et al, European patent application No. 89300963.9. Unfortunately, most of these approaches are either technically difficult to perform, expensive, or rely on the subjective evaluation of a clinician which leads to unacceptable rates of misdiagnosis.
Recently many diagnostic and forensic assays have been proposed which are based on the amplification and detection of highly polymorphic classes of repetitive DNA that are present in the human genome, e.g. Craig et al, J. Forensic Sci., Vol. 33, pgs. 1111-1126 (1988); Edwards et al, Genomics, Vol. 12, pgs. 241-253 (1992); Boerwinkle et al, Proc. Natl. Acad. Sci., Vol. 86, pgs. 212-216 (1989); Tautz, Nucleic Acids Research, Vol. 17, pgs. 6463-6471 (1989); and the like. Typically, a segment of DNA that contains the repeated sequence is amplified by polymerase chain reaction (PCR) and then sized by denaturing polyacrylamide gel electrophoresis. In regard to this approach, the so-called "short tandem repeat" or "STR" repetitive DNA has been of particular interest for diagnostic and mapping applications because of its size and genomic distribution. The length of the repeated unit in this class of DNA is typically from 2 to 6 nucleotides, and the overall length of the tandem repeat ranges from several tens to several hundreds of nucleotides making them convenient targets for PCR amplification and electrophoretic separation.
Unfortunately, the implementation of many of these detection schemes has been complicated because of imperfections in the amplification technology. For example, where amplification products must be electrophoretically separated for an assay readout, frequently secondary bands appear adjacent to a primary band, particularly when 2-nucleotide repeat sequences are amplified. This greatly complicates the quantitative analysis of the amplified product as the secondary bands of one allele may overlap the primary band(s) of other alleles.
It would be desirable to be able to detect aneuploidy using the DNA amplification approaches without the problems caused by spurious bands appearing in the electrophoretically separated amplification products.